Treatment of advanced melanoma is undergoing a revolution due to targeted therapeutics directed at BRAF(V600E) mutations and immune checkpoint blockade. However both the magnitude and durability of responses are very far from optimal (median progression on PLX4032 of ~8 months). This PPG takes a systematic, comprehensive, and interdisciplinary approach towards elucidating and overcoming this treatment resistance with combination approaches including drugs as well as immunotherapy. Our team consists of leaders in melanoma biology as well as principal investigators of key current clinical trials. Our approach builds on generating paired, patient-derived sensitive and resistant melanoma specimens and cell lines via a Shared Resource Core (Core A) using in vitro culture techniques and highly efficient xenografts in N0D/SCID/IL2RY-/- mice. These specimens, and corresponding patients, will be annotated by an extensive set of clinical and molecular criteria including candidate oncogene sequencing, genomic copy number, and PLX4720 ICSO. The same paired melanoma specimens will then undergo deep sequencing and full-genome. shRNA phenotypic screening (Project 1), to identify acquired mutations and gene dependencies associated with resistance (synthetic lethality). Validation/prioritization of hits will build on collective knowledge from ail 3 Projects. Project 2 will carry out parallel analyses of resistance mechanisms and hit validation in genetically defined mouse melanoma models, including the testing of candidates derived from Projects 1 & 3 and the incorporation of tumors with acquired resistance to BRAF inhibitors. Resistance mechanisms identified in mice will be utilized to prioritize among candidates among the human studies (Project 1). Project 3 will examine the MITF pathway which antagonizes apoptosis after BRAF(V600E) inhibitor drugs-offering drugable opportunities-and also study immune involvement which may be regulated by MITF via modulation of melanoma antigenicity. Combination therapy with BRAF(V600E) antagonists plus immune checkpoint blockade will be scrutinized using genetically defined mice and also studied in patients. The resulting molecular and functional datasets will be integrated via a specialized Bioinformatics and Biostatistics Core at the Broad Institute (Core B) to prioritize and validate hits from each platform, inform mechanisms of resistance, nominate biomarkers of sensitivity/resistance, and identify drug-able vulnerabilities for preclinical/clinical development. The predicted output of these extensively integrated Projects and Cores could not be replicated in separate initiatives, but will function within a highy collaborative, multi-disciplinary structure to accelerate the discovery of a cure.